Example embodiments of the inventive concept relate to a rapid antimicrobial susceptibility test, based on an analysis of changes in morphology and growth pattern of a microbial cell under different concentrations of various antimicrobial agents, and an automated cell image analysis system therefor. In particular, the antimicrobial susceptibility test is rapidly performed based on an analysis of changes in morphology and growth pattern of a microbial cell under different concentrations of various antimicrobial agents, and this makes it possible to obtain highly reliable test results faster by six to seven times than the standard method recommended by Clinical and Laboratory Standards Institute (CLSI).
An antimicrobial susceptibility test (AST) may be performed to estimate effects of antimicrobial agents that will be used for treatment of infectious diseases, and it may include examining whether there is susceptibility, based on a minimal inhibitory concentration (MIC).
A conventional method of testing a cell reaction to different drugs may include steps of disposing a cell in a liquid or solid medium, mixing a to-be-tested drug with the liquid medium (or disposing a paper disk, in which the drug is absorbed, on the solid medium) to react the drug with the cell, and measuring turbidity or absorbance representing an extent of a growth reaction of the cell to the drug. However, the conventional method is a statistical method of collecting information on many cells, not on a change of each cell, and thus, in order to obtain meaningful statistical results, the number of cells should be increased up to a specific value (typically, ten million per 1 ml or higher), and thus, it takes a long time to culture cells (typically, 16-24 hours). Furthermore, according to the conventional method, it is impossible to realize an observation of a change of each individual cell under a drug and an real-time observation of each motile cell real-time, and moreover, it takes a long time and great effort to test many drugs, because each drug should be injected by an individual injection process.
Furthermore, in the case where a solid medium is used for the AST method, a KB-test has a limitation on the number of samples that can be disposed on each medium plate, and thus, in order to perform an AST process for tens of different antimicrobial agents, many agar medium plates are required. Even when an automated system (e.g., VITEK system) capable of minimizing a test time is used for the AST process, a turbidity of bacteria should be increased up to a specific value, and thus, the AST process also suffers from a long test time (typically, 12 hours or more). In addition, since a test environment for the conventional method is different from a human body, the results may be largely different from the real phenomenon in the human body (Gregory G. Anderson, et al. (2003), “Intracellular Bacterial Biofilm-Like Pods in Urinary Tract Infections”, Science 301, 105; Gallo et al. (2011), “Demonstration of Bacillus cereus in Orthopaedic-Implant-Related Infection with Use of a Multi-Primer Polymerase Chain Reaction-Mass Spectrometric Assay”, J Bone Joint Surg Am, 93).
To overcome technical limitations of the conventional AST method, there have been proposed various methods, called rapid AST (RAST), which make it possible to observe the division of microbes at an initial step. For example, each of such known methods may include a step of measuring the number of microbial cells in micro-fluidic channel, measuring a rotation speed of a magnetic bead to estimate a weight of microbes, measuring a fluorescence signal associated with metabolic activity of microbial cells in liquid droplet, or calculating an area of an image, which is occupied by microbes, to estimate susceptibility to an antimicrobial agent.
Meanwhile, microbial reactions to antimicrobial agents are very heterogeneous and are specifically dependent on antimicrobial agent conditions, but the known methods are performed based on the observation for examining whether a microbe is growing. In other words, in the known methods, only the presence or absence of ‘growth’ of a microbial cell is observed, without consideration for various morphological changes (other than ‘growth’ or ‘no-growth’) of a microbial cell, which may occur under different conditions of antimicrobial agents, and thus, the known methods suffer from low reliability of AST results.